前列腺素I2合成酶调节巨噬细胞极化并促进结直肠癌淋巴结转移研究  

Prostaglandin I2 synthase regulates macrophage polarization and promotes lymph node metastasis in colorectal cancer

作　　者：程华 郑勇斌[1] Cheng Hua;Zheng Yongbin(Department of Gastrointestinal Surgery,Renmin Hospital of Wuhan University,Wuhan 430060,China)

机构地区：[1]武汉大学人民医院胃肠外科,武汉430060

出　　处：《中华实验外科杂志》2024年第3期445-448,共4页Chinese Journal of Experimental Surgery

基　　金：湖北省陈孝平科技发展基金会(CXPJJH121003-2103)。

摘　　要：目的探究脂代谢基因在结直肠癌(CRC)淋巴结转移中的生物学作用及其机制。方法选取8例患者淋巴结转移和未转移CRC组织进行高通量测序,结合公共数据库筛选与淋巴结转移及预后相关的脂代谢基因,即前列腺素I2合成酶(PTGIS)。在收集的70例CRC组织中验证PTGIS表达与淋巴结转移的关系。接下来使用小干扰RNA(siRNA)在CRC细胞系中沉默PTGIS,将细胞分为转染组(siPTGIS)和对照组,荧光实时定量聚合酶链反应(RT-qPCR)及蛋白质印迹法(Western blot)验证沉默效率。采用细胞计数盒(CCK-8)、单克隆实验和5-乙炔基-2’-脱氧尿苷(EdU)染色分析沉默PTGIS对CRC增殖能力的影响;采用Transwell实验分析PTGIS沉默对CRC细胞迁移和侵袭能力影响;采用尼罗绿染色和甘油三酯定量试剂盒检测细胞的脂质水平;采用流式细胞仪分析沉默PTGIS的CRC细胞与THP-1共培养24 h后巨噬细胞表型变化。采用免疫荧光(IF)验证PTGIS与M1型巨噬细胞表达丰度的关系,两组间比较采用t检验。结果免疫组织化学(IHC)显示淋巴结转移的CRC组织中PTGIS的表达水平高于无淋巴结转移CRC组织(25.74±1.36比5.10±0.57,t=13.96,P<0.01);PTGIS沉默后CRC细胞的增殖、迁移及侵袭能力显著低于对照组(HCT8:CCK-8:0.64±0.01比1.06±0.01,t=22.31,P<0.01;EdU:35.20±0.92比64.70±1.62,t=15.75,P<0.01;单克隆实验:761.00±46.36比1672.00±166.70,t=5.26,P<0.01;Transwell迁移:53.00±1.73比92.67±1.20,t=18.82,P<0.01;Transwell侵袭:27.67±1.20比65.00±2.08,t=15.53,P<0.01);转染组中脂滴数目和甘油三酯含量明显低于对照组(脂滴:0.39±0.01比1.24±0.07,t=10.41,P<0.01;甘油三酯:74.50±1.50比142.50±1.50,t=32.06,P<0.01);转染组的CRC细胞与THP-1共培养后M1型巨噬细胞比例高于对照组(1.86±0.06比1.00±0.00,t=13.97,P<0.01);PTGIS低表达CRC组织中M1型巨噬细胞表达高于PTGIS高表达CRC组织(1.17±0.01比0.66±0.02,t=23.12,P<0.01)。结论沉默PTGIS抑制CRC细胞转移并促进巨Objective To explore the biological role of lipid metabolism genes in colorectal cancer(CRC)lymph node metastasis and its mechanisms.MethodsFirst,8 cases of lymph node metastasis and non-metastatic CRC tissues were selected for high throughput sequencing,and public databases were combined to screen for lipid metabolism genes related to lymph node metastasis and prognosis,namely prostaglandin I2 synthase(PTGIS).Subsequently,the relationship between PTGIS expression and lymph node metastasis was verified in 70 CRC tissues.Next,PTGIS was silenced in CRC cell lines using small interfering RNA(siRNA),the cells were divided into transfected(siPTGIS)and control groups,and silencing efficiency was verified by real-time fluorescent quantitative polymerase chain reaction(RT-qPCR)and Western blotting.Cell counting kit-8(CCK-8)assay,monoclonal assay and 5-ethynyl-2-deoxyuridine(EdU)staining were used to analyze the effect of silencing PTGIS on the proliferative ability of CRC.Transwell assay was used to analyze the effect of PTGIS silencing on the migratory and invasive ability of CRC cells.Nile green staining and Triglyceride quantification kit were used to detect the lipid level of the cells.Flow cytometry was used to analyze the effect of silencing PTGIS on the macrophage phenotypic changes of CRC cells co-cultured with THP-1 after 24 h of co-culture with THP.Finally,immunofluorescence(IF)was used to verify the relationship between PTGIS and the expression abundance of M1-type macrophages.T-test was used for comparisons between groups.ResultsImmunohistochemistry(IHC)showed higher expression levels of PTGIS in CRC tissues with lymph node metastasis than in CRC tissues without lymph node metastasis(25.74±1.36 vs.5.10±0.57,t=13.96,P<0.01).The proliferation,migration and invasion ability of CRC cells after PTGIS silencing was significantly lower than that of the control group(HCT8:CCK-8:0.64±0.01 vs.1.06±0.01,t=22.31,P<0.01;EdU:35.20±0.92 vs.64.70±1.62,t=15.75,P<0.01;monoclonal assay:761.00±46.36 vs.1672.00±166.70,t=5

关 键 词：前列腺素I2合成酶 结直肠癌 淋巴结转移 脂代谢重编程 巨噬细胞极化 

分 类 号：R735.34[医药卫生—肿瘤]